1. Field of the Invention
Compounds that modulate water transport by aquaporin channels in cell membranes and methods of using these compound to treat diseases and disorders associated with aquaporin activity.
2. Description of the Related Art
Aquaporins (AQPs) represent a diverse family of membrane proteins found in prokaryotes and eukaryotes (Reizer et al., 1993; Hohmann et al., 2000; King et al., 2004). Aquaporin channels have a well known role for promoting transmembrane diffusion of water (Agre et al., 1993), as well a other molecules including ions, gases, and small organic compounds (Yool and Stamer, 2002). Aquaporins are encoded by members of the major intrinsic protein (MIP) gene family. In mammals, there are at least 12 classes of aquaporins (AQP0 to AQP11) that exhibit tissue-specific patterns of expression (Nielsen et al., 2002). Aquaporin channels play roles in various physiological processs and have been implicated in pathophysiological conditions involving water imbalance.
AQP1 is a widely expressed water channel found in the basolateral and apical plasma membranes of the proximal tubules, the descending limb of the loop of Henle, and in the descending portion of the vasa recta of the kidney. It is also present in red blood cells, vascular endothelium, the gastrointestinal tract, sweat glands, and lungs.
AQP4 is expressed abundantly throughout the central nervous system, where it is concentrated in the perivascular endfeet of astroglial cells that surround blood vessels and maintain the integrity of the blood-brain barrier, suggesting an important role in the regulation of brain water balance (Nagelhus et al., 1998; Rash et al., 1998). Colocalization of AQP4 in astroglial endfeet occurs with Kir1.4 (inward rectifier) and Kca (Ca+2-activated) potassium channels. AQP4 may be part of a complex organization of proteins that control salt and water influx (Price et al., 2002). Activation of protein kinase C (PKC) stimulates phosphorylation of AQP4 at serine 180 (Zelenina et al., 2002), and causes dose-dependent inhibition of water permeability. Levels of AQP4 expression also are influenced by protein kinase C (Yamamoto et al., 2001; (Han et al., 1998)), bradykinin (Monti et al., 2001), vasopressin (Niermann et al., 2001), and ischemia-related genes (Nicchia et al., 2003), and are massively upregulated in tumor-associated reactive astrocytes (Saadoun et al., 2002).
Verkman and colleagues (Manley et al., 2000) compared the effects of water intoxication and permanent stroke in wild type (+/+) and AQP4 knockout (−/−) mice. Water intoxication was imposed by giving mice intraperitoneal injections of distilled water equal to 20% of body weight. Outcomes focused on mortality, observed neurological deficits, and electron microscopic analyses of brain ultrastructure. Astrocytic endfoot areas in ten randomly selected micrographs were measured from wild-type and AQP4 deficient mice. The lower water content in brain tissue and the reduced astrocytic swelling in the AQP4−/− mice were consistent with improved neurological deficit scores and substantially better survival as compared with the AQP4+/+ mice.
Permanent middle cerebral artery (MCA) occlusion also was tested in AQP+/+ and AQP4−/− mice (n=10 per group). The measured outcomes at 24 h were hemisphere size and neurological deficits (FIG. 1). Significantly greater hemispheric enlargement from brain edema occurred in wild type (FIG. 1A). AQP4+/+ mice also had significantly higher (worse) neurological deficit scores as compared with the AQP4−/− mice (FIG. 1B). Manley and colleagues suggested that the reduced brain edema in AQP4-deficient mice indicates “the AQP4 water channel is a potential target for drug discovery.”
AQP9 may also influence brain edema. It is permeable to water and small solutes (urea, glycerol), and is expressed in brain glial cells as well as in liver, spleen and other organs (Badaut et al., 2001; Badaut et al., 2002; Elkjaer et al., 2000; Nicchia et al., 2001). AQP9 is present in cell bodies and processes of astrocytes in contact with brain vessels and fluid compartments. In contrast with AQP4, AQP9 staining is not polarized on astrocytic endfeet (Badaut et al., 2004).
Edema results from many clinical conditions including several neurological conditions or pulmonary conditions. A major unmet medical need is treatment of edema associated with stroke. Stroke, as the third leading cause of death, is a devastating disease affecting more than 700,000 people in the United States each year. The extent of brain edema is a major determinant of patient survival after a stroke event (Dirnagl et al., 1999; Taylor et al., 1996). For progressive edema due to middle cerebral artery occlusion, mortality approaches 80% (Ayata and Ropper, 2002). The propensity of ischemic brain tissue to develop edema remains the major cause of death in patients with large infarctions, particularly within the middle cerebral artery territory and cerebellum involved in 15-20% of all strokes (Hacke et al., 1996). An important problem recognized in the prior art was the identification of molecular targets for intervention in the edema process. However, clinically acceptable strategies for management of ischemic brain edema have remained elusive, and available treatments are often of limited value for patients with massive edema.
Brain edema is classified as vasogenic (movement of water and solutes across the blood brain barrier), or cytotoxic (osmotic swelling of cells in the affected area) (Fishman, 1975; Klatzo, 1967). Cytotoxic edema correlates with initial infarct size, and vasogenic edema contributes to the delayed risk-prone processes of brain swelling (FIG. 2). Astrocytes play a role in both processes. Astrocyte endfeet covering the abluminal capillary surface promote maintenance of intercellular tight junctions between endothelial cells that create the blood-brain barrier (Huber et al., 2001). Blood-brain barrier disruption after ischemia involves altered transport, opening of paracellular pathways, and physical disruption of astrocyte-endothelial junctions (Sun and O'Donnell, 1996; Venero et al., 2001). Pericapillary astrocyte endfeet are the first cellular elements to swell during brain ischemia (Dodson et al., 1977), a process thought to result from uptake of extracellular K+, Cl−, and Na+ and the osmotic flux of water (Su et al., 2002), promoting a progressive cascade of pathology.
Aquaporin expression has been associated with the formation of brain edema in studies comparing edema between mice expressing AQP4 and knockout mice not expressing AQP4. Less hemispheric enlargement was observed in knock out mice (FIG. 1A) and neurological scores of knockout mice improved (FIG. 1B), see (Manley et al., 2004). In addition, the selective removal of perivascular AQP4 by β-syntrophin deletion has been shown to delay the buildup of brain edema (assessed by Diffusion-weighted MRI) following water intoxication, despite the presence of a normal complement of endothelial AQP4 (Amiry-Moghaddam et al., 2006).
Moreover, a significant positive correlation between AQP4 mRNA expression and blood brain barrier (BBB) permeability after experimental cerebral hemorrhage in rats has been observed and these changes are consistent with observed increases in cerebral edema. Other results associate the expression of AQP4 with edema and blood brain permeability (Teng et al., 2006; Chen et al., 2006). Inflammation related edema is also associated with AQP4. Cysteinyl leukotrienes (including LTC4, LTD4, and LTE4) are potent inflammatory mediators that can induce brain-blood barrier (BBB) disruption and brain edema. LTD4 has been shown to affect brain edema in two ways; the CysLT1 receptor mediates vasogenic edema while CysLT2 receptor may mediate cytotoxic edema via up-regulating AQP4 expression (Wang et al., 2006).
Brain edema is associated with a variety of infectious diseases of the brain. In one study, Streptococcus pneumoniae was injected into cerebrospinal fluid (CSF) in wild type and AQP4 null mice. AQP4 protein was found to be strongly up-regulated as a result of meningitis, with an approximately 5-fold increase in water permeability (Pf) across the blood-brain barrier observed compared to non-infected wild type mice (Papadopoulos, et al., Journal of Biological Chemistry (2005), 280(14), 13906-13912. Conversely, other have demonstrated a protective effect of AQP4 on brain swelling in bacterial abscess, suggesting that AQP4 induction may reduce vasogenic edema associated with cerebral infection (Bloch, et al., Journal of Neurochemistry (2005), 95(1), 254-262).
The expression of mRNA for AQP1 and AQP4 was analyzed in a well-established mice model that simulates encephalitis caused by the human Herpes simplex virus encephalitis (HSVE). This study is consistent with a significant down-regulation of AQP4 in the acute phase of disease and an up-regulation of AQP1 and AQP4 in the long term. AQP4 modulation has been suggested as a potential target for treating brain edema associated with herpetic infection by HSVE (Tones et al., Journal of NeuroVirology (2007), 13(1), 38-46).
Two members of the aryl-sulfonamide structural class have been reported as inhibitors of the AQP4 channel (Huber et al., Biorganic & Medicianal Chemistry Letters (2007), 17, 1270-1273). One of these compounds, acetazolamde, has recently been claimed as a carbonic anhydrase inhibitor for the treatment of unstable cerebralspinal pressure syndrome (Yoshida, Application; WO 2006-JP322065 20061030). However, there is a lack of reliable information about the structures of compounds that regulate AQP4 function (Dun et al., Neural Regeneration Research (2007), 2(4), 234-238).
It has been an unachieved objective to identify and characterize aquaporin modulators, such as AQP4 blockers or activators, as a potential adjunct therapy for clinical intervention in edema and stroke-related pathologies (Zador et al., Progress in brain research (2007), 161 185-94).
While little has previously been known regarding the pharmacology of AQP channels, some compounds, such as mercury and organic mercurials, selectively block some types of AQPs, such as AQP1, but not AQP4. However, these blockers exhibit high systemic toxicity negatating their potential therapeutic value. Other aquaporin blockers, such as tetraethylammonium and phloretin suffer from a lack of specificity and potency. While AQP4 expression have been associated with phenomena like edema, suitable pharmacological compounds for modulating AQP4 activity or suitable for prospective clinical use have not been available. There is also a significant need to develop novel and/or selective AQP modulators for further investigation of the role of aquaporins in edema and other disease phenomena.
To address these problems, the inventors have identified novel compounds that agonize or antagonize aquaporin activity and which lack the toxicity, poor potency and low specificity of prior art compounds.